Rapid Mutation of Endogenous Zebrafish Genes Using Zinc Finger Nucleases Made by Oligomerized Pool ENgineering (OPEN)

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Rapid Mutation of Endogenous Zebrafish Genes Using Zinc Finger Nucleases Made by Oligomerized Pool ENgineering (OPEN)

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dc.contributor.author Foley, Jonathan E.
dc.contributor.author Reyon, Deepak
dc.contributor.author Yeh, Jing-Ruey Joanna
dc.contributor.author Maeder, Morgan L.
dc.contributor.author Sander, Jeffry D
dc.contributor.author Peterson, Randall Theodore
dc.contributor.author Joung, Jae Keith
dc.date.accessioned 2011-01-26T12:58:25Z
dc.date.issued 2009
dc.identifier.citation Foley, Jonathan E., Jing-Ruey J. Yeh, Morgan L. Maeder, Deepak Reyon, Jeffry D. Sander, Randall T. Peterson, and J. Keith Joung. 2009. Rapid mutation of endogenous zebrafish genes using zinc finger nucleases made by Oligomerized Pool ENgineering (OPEN). PLoS ONE 4(2): e4348. en_US
dc.identifier.issn 1932-6203 en_US
dc.identifier.uri http://nrs.harvard.edu/urn-3:HUL.InstRepos:4687054
dc.description.abstract Background: Customized zinc finger nucleases (ZFNs) form the basis of a broadly applicable tool for highly efficient genome modification. ZFNs are artificial restriction endonucleases consisting of a non-specific nuclease domain fused to a zinc finger array which can be engineered to recognize specific DNA sequences of interest. Recent proof-of-principle experiments have shown that targeted knockout mutations can be efficiently generated in endogenous zebrafish genes via non-homologous end-joining-mediated repair of ZFN-induced DNA double-stranded breaks. The Zinc Finger Consortium, a group of academic laboratories committed to the development of engineered zinc finger technology, recently described the first rapid, highly effective, and publicly available method for engineering zinc finger arrays. The Consortium has previously used this new method (known as OPEN for Oligomerized Pool ENgineering) to generate high quality ZFN pairs that function in human and plant cells. Methodology/Principal Findings: Here we show that OPEN can also be used to generate ZFNs that function efficiently in zebrafish. Using OPEN, we successfully engineered ZFN pairs for five endogenous zebrafish genes: tfr2, dopamine transporter, telomerase, hif1aa, and gridlock. Each of these ZFN pairs induces targeted insertions and deletions with high efficiency at its endogenous gene target in somatic zebrafish cells. In addition, these mutations are transmitted through the germline with sufficiently high frequency such that only a small number of fish need to be screened to identify founders. Finally, in silico analysis demonstrates that one or more potential OPEN ZFN sites can be found within the first three coding exons of more than 25,000 different endogenous zebrafish gene transcripts. Conclusions and Significance: In summary, our study nearly triples the total number of endogenous zebrafish genes successfully modified using ZFNs (from three to eight) and suggests that OPEN provides a reliable method for introducing targeted mutations in nearly any zebrafish gene of interest. en_US
dc.language.iso en_US en_US
dc.publisher Public Library of Science en_US
dc.relation.isversionof doi:10.1371/journal.pone.0004348 en_US
dc.relation.hasversion http://www.ncbi.nlm.nih.gov/pmc/articles/PMC2634973/pdf/ en_US
dash.license LAA
dc.title Rapid Mutation of Endogenous Zebrafish Genes Using Zinc Finger Nucleases Made by Oligomerized Pool ENgineering (OPEN) en_US
dc.type Journal Article en_US
dc.description.version Version of Record en_US
dc.relation.journal PLoS ONE en_US
dash.depositing.author Joung, Jae Keith
dc.date.available 2011-01-26T12:58:25Z
dash.affiliation.other HMS^Medicine-Massachusetts General Hospital en_US
dash.affiliation.other HMS^Pathology en_US

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